*Article De Novo* **Transcriptome Meta-Assembly of the Mixotrophic Freshwater Microalga** *Euglena gracilis*

**Javier Cordoba <sup>1</sup> , Emilie Perez 1,2, Mick Van Vlierberghe <sup>2</sup> , Amandine R. Bertrand <sup>2</sup> , Valérian Lupo <sup>2</sup> , Pierre Cardol <sup>1</sup> and Denis Baurain 2,\***


**Abstract:** *Euglena gracilis* is a well-known photosynthetic microeukaryote considered as the product of a secondary endosymbiosis between a green alga and a phagotrophic unicellular belonging to the same eukaryotic phylum as the parasitic trypanosomatids. As its nuclear genome has proven difficult to sequence, reliable transcriptomes are important for functional studies. In this work, we assembled a new consensus transcriptome by combining sequencing reads from five independent studies. Based on a detailed comparison with two previously released transcriptomes, our consensus transcriptome appears to be the most complete so far. Remapping the reads on it allowed us to compare the expression of the transcripts across multiple culture conditions at once and to infer a functionally annotated network of co-expressed genes. Although the emergence of meaningful gene clusters indicates that some biological signal lies in gene expression levels, our analyses confirm that gene regulation in euglenozoans is not primarily controlled at the transcriptional level. Regarding the origin of *E. gracilis*, we observe a heavily mixed gene ancestry, as previously reported, and rule out sequence contamination as a possible explanation for these observations. Instead, they indicate that this complex alga has evolved through a convoluted process involving much more than two partners.

**Keywords:** transcriptome assembly; gene expression; transcriptional regulation; ontology network; co-expression network; taxonomic analysis; database contamination; kleptoplastidy
